Abstract-The formation of advanced glycation end products (AGEs) on extracellular matrix components leads to accelerated increases in collagen cross linking that contributes to myocardial stiffness in diabetes. This study determined the effect of the crosslink breaker, ALT-711 on diabetes-induced cardiac disease.

The renin-angiotensin system (RAS) has an important role in the endocrine pancreas. Although angiotensin II has significant effects on cell proliferation and apoptosis, the contribution of the RAS to changes in islet structure and function associated with type 2 diabetes is yet to be defined. This study examined the specific effects of RAS blockade on islet structure and function in diabetes. Thirty-six male Zucker diabetic fatty (ZDF) rats, 10 weeks of age, were randomized to receive the angiotensin-converting enzyme inhibitor perindopril (8 mg/l in drinking water; n ‫؍‬ 12), irbesartan (15 mg/kg via gavage; n ‫؍‬ 12), or no treatment (n ‫؍‬ 12) for 10 weeks. Results were compared with lean littermates (ZL) (n ‫؍‬ 12) studied concurrently. ZDF rats had increased intraislet expression of components of the RAS correlating with increased intraislet fibrosis, apoptosis, and oxidative stress. Disordered islet architecture, seen in ZDF rats, was attenuated after treatment with perindopril or irbesartan. Islet fibrogenesis was also diminished, as measured by picrosirius staining and expression of collagens I and IV. Gene expression of transforming growth factor-␤1 was increased in the ZDF pancreas (ZL, 1.0 ؎ 0.1; ZDF, 2.0 ؎ 0.3; P < 0.05) and reduced after blockade of the RAS (ZDF ؉ P, 1.3 ؎ 0.2; ZDF ؉ I, 1.5 ؎ 0.1; vs. ZDF, both P < 0.05). Improvements in structural parameters were also associated with functional improvements in first-phase insulin secretion. These findings provide a possible mechanism for the reduced incidence of new-onset diabetes that has been observed in clinical trials of RAS blockade. Diabetes 53: 989 -997, 2004

Background-Atherosclerosis is a major complication of diabetes, but the mechanisms by which diabetes promotes macrovascular disease have not been fully delineated. Although several animal studies have demonstrated that inhibition of ACE results in a decrease in the development of atherosclerotic lesions, information about the potential benefits of these agents on complex and advanced atherosclerotic lesions as observed in long-term diabetes is lacking. The aim of this study was to evaluate whether treatment with the ACE inhibitor perindopril affects diabetes-induced plaque formation in the apolipoprotein E (apoE)-deficient mouse. Methods and Results-Diabetes was induced by injection of streptozotocin in 6-week-old apoE-deficient mice. Diabetic animals received treatment with perindopril (4 mg · kg Ϫ1 · d

Advanced glycation end product (AGE) formation may contribute to the progression of atherosclerosis, particularly in diabetes. The present study explored atherosclerosis in streptozotocin-induced diabetic apolipoprotein E-deficient (apoE؊/؊) mice that were randomized (n ‫؍‬ 20) to receive for 20 weeks no treatment, the AGE cross-link breaker ALT-711, or the inhibitor of AGE formation aminoguanidine (AG). A sixfold increase in plaque area with diabetes was attenuated by 30% with ALT-711 and by 40% in AG-treated mice. Regional distribution of plaque demonstrated no reduction in plaque area or complexity within the aortic arch with treatment, in contrast to the thoracic and abdominal aortas, where significant attenuation was seen. Diabetes-associated accumulation of AGEs in aortas and plasma and decreases in skin collagen solubility were ameliorated by both treatments, in addition to reductions in the vascular receptor for AGE. Collagenassociated reductions in the AGEs carboxymethyllysine and carboxyethyllysine were identified with both treatments. Diabetes was also accompanied by aortic accumulation of total collagen, specifically collagens I, III, and IV, as well as increases in the profibrotic cytokines transforming growth factor-␤ and connective tissue growth factor and in cellular ␣-smooth muscle actin. Attenuation of these changes was seen in both treated diabetic groups. ALT-711 and AG demonstrated the ability to reduce vascular AGE accumulation in addition to attenuating atherosclerosis in these diabetic mice.

Background-It remains controversial whether specific blockade of the renin-angiotensin system confers superior antiatherosclerotic effects over other antihypertensive agents in diabetes. Therefore, the aim of this study was to compare equihypotensive doses of the angiotensin II subtype 1 (AT 1 ) receptor blocker irbesartan with the calcium antagonist amlodipine on diabetes-induced plaque formation in the apolipoprotein E (apoE)-null mouse and to explore molecular and cellular mechanisms linked to vascular protection. Methods and Results-Diabetes was induced by injection of streptozotocin in 6-week-old apoE-null mice. Diabetic animals were randomized to no treatment, irbesartan, or amlodipine for 20 weeks. Diabetes was associated with an increase in plaque area and complexity in the aorta in association with a significant increase in aortic AT 1 receptor expression, cellular proliferation, collagen content, macrophage-and ␣-smooth muscle actin-positive cell infiltration, as well as an increased expression of platelet-derived growth factor-B (PDGF-B), monocyte chemoattractant protein-1 (MCP-1), and vascular cell adhesion molecule-1 (VCAM-1). Irbesartan but not amlodipine treatment attenuated the development of atherosclerosis, collagen content, cellular proliferation, and macrophage infiltration as well as diabetes-induced AT 1 receptor, PDGF-B, MCP-1, and VCAM-1 overexpression in the aorta despite similar blood pressure reductions by both treatments. Conclusions-Diabetes-associated atherosclerosis is ameliorated by AT 1 receptor blockade but not by calcium channel antagonism, providing further evidence for the vascular renin-angiotensin system playing a pivotal role in the development and acceleration of atherosclerosis in diabetes.

Background-Although in vitro studies have suggested that tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) might be involved in vascular biology, its potential role in the pathogenesis and/or treatment of atherosclerosis has not been investigated. Methods and Results-Both recombinant human TRAIL and an adeno-associated virus vector expressing human TRAIL were used to deliver TRAIL in apolipoprotein E (apoE)-null mice in which diabetes mellitus was induced by destruction of islet cells with streptozotocin. Diabetes in apoE-null mice was associated with a significant increase in atherosclerotic plaque area and complexity in the aorta as assessed by a marked increase in interstitial collagen, cellular proliferation, and macrophage infiltration and a focal loss of endothelial coverage. Repeated intraperitoneal injections of recombinant human TRAIL and a single intravenous injection of adeno-associated virus-human TRAIL significantly attenuated the development of atherosclerotic plaques in apoE-null animals. TRAIL also markedly affected the cellular composition of plaque lesions by inducing apoptosis of infiltrating macrophages and increasing the vascular smooth muscle cell content. Moreover, TRAIL promoted the in vitro migration of cultured human aortic vascular smooth muscle cells but not of monocytes or macrophages. Conversely, TRAIL selectively induced apoptosis of human cultured macrophages but not of vascular smooth muscle cells. Conclusions-Overall, data from the present study indicate that atherosclerosis in diabetic apoE-null mice is ameliorated by systemic TRAIL administration and that adeno-associated virus-mediated TRAIL gene delivery might represent an innovative method for the therapy of diabetic vascular diseases.

Abstract. Hyperlipidemia not only may be relevant to cardiovascular disease in diabetes but may also play a role in the development and progression of diabetic nephropathy. Furthermore, there is increasing evidence that advanced glycation end products (AGE) play an important role in diabetic renal disease. The objectives of this study were first to characterize renal injury in diabetic apolipoprotein E knockout (apo E-KO) mice and second to explore the role of AGE in the development and progression of renal disease in this model. Diabetes was induced by injection of streptozotocin in 6-wk-old apo E-KO mice. Diabetic animals received no treatment or treatment with the inhibitor of AGE formation aminoguanidine (1 g/kg per d) or the cross-link breaker [4,5-dimethyl-3-(2-oxo2-phenylethyl)-thiazolium chloride] ALT-711, which cleaves preformed AGE (20 mg/kg per d) for 20 wk. Nondiabetic apo E-KO mice as well as nondiabetic and diabetic C57BL/6 mice served as controls. Compared with nondiabetic apo E-KO mice, induction of diabetes in apo E-KO mice resulted in accelerated renal injury characterized by albuminuria and glomerular and tubulointerstitial injury. These abnormalities were associated with increased expression of collagen type I and type IV and transforming growth factor-␤1 (TGF-␤1), increased ␣-smooth muscle actin immunostaining and macrophage infiltration, and increased serum and renal AGE. The two treatments, which attenuated renal AGE accumulation in a disparate manner, were associated with less albuminuria, structural injury, macrophage infiltration, TGF-␤1, and collagen expression. The accelerated renal injury that was observed in diabetic apo E-KO mice was attenuated by approaches that inhibit renal AGE accumulation.

S U M M A R YThe temporal and spatial expression of transforming growth factor (TGF)-␤ 1 and connective tissue growth factor (CTGF) was assessed in the left ventricle of a myocardial infarction (MI) model of injury with and without angiotensin-converting enzyme (ACE) inhibition. Coronary artery ligated rats were killed 1, 3, 7, 28, and 180 days after MI. TGF-␤ 1 , CTGF, and procollagen ␣ 1(I) mRNA were localized by in situ hybridization, and TGF-␤ 1 and CTGF protein levels by immunohistochemistry. Collagen protein was measured using picrosirius red staining. In a separate group, rats were treated for 6 months with an ACE inhibitor. There were temporal and regional differences in the expression of TGF-␤ 1 , CTGF, and collagen after MI. Procollagen ␣ 1(I) mRNA expression increased in the border zone and scar peaking 1 week after MI, whereas collagen protein increased in all areas of the heart over the 180 days. Expression of TGF-␤ 1 mRNA and protein showed major increases in the border zone and scar peaking 1 week after MI. The major increases in CTGF mRNA and protein occurred in the viable myocardium at 180 days after MI. Long-term ACE inhibition reduced left ventricular mass and decreased fibrosis in the viable myocardium, but had no effect on cardiac TGF-␤ 1 or CTGF. TGF-␤ 1 is involved in the initial, acute phase of inflammation and repair after MI, whereas CTGF is involved in the ongoing fibrosis of the heart. The antifibrotic benefits of captopril are not mediated through a reduction in CTGF.